Lasse Greiner et al.
COMMUNICATIONS
with an uncoated fused silica capillary (CElectFS50,
SUPELCO, length to detection window 43 cm, total length
50 cm in a 40 mmol LÀ1 potassium phosphate and 10 mmol LÀ1
borate buffer at pH 8.5, 408C, 30 kV, migration times:
metal block. The total volume was increased to 150 mL. For
batch operation the ultrafiltration unit was omitted from the
set-up. Prior to reaction, the reactor was flushed with a solution
of 25 mol LÀ1 AAZ in methanol (1 L) for ten hours. During this
time the volume aeration unit was flushed with hydrogen at
2 mL minÀ1. The PyrPhos catalyst was dissolved in 5 mL
methanol and added via the dosing pump (resulting concen-
tration 1 mmol LÀ1). Throughout the reaction a flow of
1 mL hÀ1 was maintained, and the outflow was analyzed by
capillary electrophoresis. The hydrogen flow to maintain
constant pressure on the gaseous side was measured by mass
flow metering (Bronkhorst, Ruurlo, The Netherlands), and
recorded on a personal computer.
uridine (internal standard) 2.9 min, NADP 9.1 min, NADPH
12.1 min. Protein content was determined by the method
described by Sedmark et al.,[14] by measuring the absorption of
the Coomassie Brilliant Blue G250 complex (l 620 nm)
against samples of known concentration of bovine serum
albumin. Hydrogenase was stored in liquid nitrogen and
thawed directly before use under an argon atmosphere, which
was replaced with hydrogen before heating to 808C for 10 min.
Volume-Aerated Membrane Reactor
Acknowledgements
The reactor set-up is depicted in Figure 1. All connections were
made using metal fittings (Swagelok, Neufahrn, Germany) or if
not possible polymer fittings (Upchurch Scientific, Oak
Harbor, USA). The dosage pump was an alternating piston
pump (P500, Pharmacia, Erlangen, Germany) controlled via a
personal computer using the signal of the mass flow meter
(Bronkhorst, Ruurlo, The Netherlands), as described else-
where.[2] Circulation flow in the aeration unit was established
by a magnet coupled gear pump (Verder, Haan, Germany).
The membrane filtration unit was a modified set-up used
before[2] equipped with an ultrafiltration membrane (Amicon
YM10, Millipore, Eschborn, Germany) and it was at the same
time used as heat exchanger. The volume aeration unit was a
The authors would like to thank HuubHaaker, Hans Waasink
(University of Wageningen, Netherlands) and Colja Laane
(DSM, Netherlands) for discussions and the kind gift of
hydrogenase preparation; Dietmar Reichert (Degussa, Germa-
ny) for the gift of PyrPhos catalyst and discussions. Further-
more, thanks are due to Degussa and the German Federal
Ministry of Education and Research for financial support
concerning the chemical catalysis.
6 mm metal tube of 35 cm length. In this tube the PTFE References
membrane (CS-Chromatographie Service, Langerwehe, Ger-
many) of 1.6 mm outer and 1.0 mm inner diameter was
concentrically positioned. Pressure was measured by pressure
transmitters (Wika, Klingenberg, Germany). Both liquid and
gas pressure were controlled via membrane valves (gas: AGA,
Hamburg, Germany; liquid: Jasco, Gross-Umstadt, Germany).
The technical data of the set-up is given in Table 1.
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Continuous Operation of the Volume-Aerated
Membrane Reactor: Enzyme Catalysis
Prior to addition of the catalyst, the reactor was flushed with
degassed buffer for at least 5 residence times (reactor volume
25 mL), subsequently with 50 mL of substrate solution. During
this time the volume aeration unit was flushed with hydrogen at
2 mL minÀ1. The reactor temperature was maintained at 408C
by a thermostatic water bath (Lauda, Lauda-Kˆnigshofen,
Germany). Prior to addition of the enzyme, 100 mg of bovine
serum albumin was added to the reactor via the dosing pump,
and the enzyme preparation was heated under hydrogen
atmosphere for 10 min before adding it to the reactor.[13] The
course of the reaction and the other conditions are given in
Figure 2. Samples of the outflow were analyzed.
Batch Operation of the Volume-Aerated Reactor:
Chemical Catalysis
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The aeration unit was replaced by another one with a larger
surface area of 400 cm2. The 10 m of PTFE tubing of the same
dimensions used above were coiled onto a hexagonal inner
682
¹ 2003 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
asc.wiley-vch.de
Adv. Synth. Catal. 2003, 345, 679 683